Hepatitis C virus (HCV) infection is a major world health problem infecting 170 million people with many chronic HCV carriers at risk of developing liver cirrhosis and liver cancer. There is a strong correlation between chronic HCV infection, low plasma cholesterol levels and steatosis (fatty liver). Little is known about the role of HCV in the pathogenesis of steatosis. Understanding the mechanism of steatosis associated with hepatitis C is relevant to the design of antiviral therapies. In this study, we propose to test the hypothesis that HCV-induced oxidative stress triggers intracellular events, which lead to hepatosteatosis and fibrosis. This hypothesis is strengthened by our recent preliminary results in which we observed accumulation of cholesterol in HCV genotype 1-expressing cells. The specific aims of this proposal are to: (1) to investigate the mechanism(s) of activation of sterol regulatory element binding proteins (SREBPs) by HCV gene expression and (2) to analyze the effect of HCV gene expression on the major components of the cholesterol and lipid biosynthesis pathways by determining the direct affect of HCV gene expression on the proteins and enzymes involved in cholesterol and lipid biosynthesis. SREBPs are endoplasmic reticulum (ER)-associated proteins, which are cleaved upon sensing low sterols. The liberated proteins then translocate to the nucleus and stimulate genes involved in lipid and fatty acid synthetic pathways. Using the HCV replicon bearing cells, we propose to monitor the activation of SREBP proteins. Using the system we furthe propose to determine the effects of HCV gene expression on the regulation of microsomal trigylceride protein (MTP) and lipid export. To correlate with projects of the center grant, the role of HCV gene expression in altering MTP function and its relevance to CD1d cell surface expression will be investigated. Finally, the conditioned culture supernatants will be tested for their fibrogenic potential via cytokine signaling. These collaborative efforts provide a coherent theme of the center and are likely to offer insight into the mechanisms of liver injury including steatosis associated with chronic HCV infection as well as guide the design of novel therapeutic approaches.